Signaling – transcription interactions in mouse retinal ganglion cells early axon pathfinding –a literature review

Sending an axon out of the eye and into the target brain nuclei is the defining feature of retinal ganglion cells (RGCs). The literature on RGC axon pathfinding is vast, but it focuses mostly on decision making events such as midline crossing at the optic chiasm or retinotopic mapping at the target nuclei. In comparison, the exit of RGC axons out of the eye is much less explored. The first checkpoint on the RGC axons’ path is the optic cup - optic stalk junction (OC-OS). OC-OS development and the exit of the RGC pioneer axons out of the eye are coordinated spatially and temporally. By the time the optic nerve head domain is specified, the optic fissure margins are in contact and the fusion process is ongoing, the first RGCs are born in its proximity and send pioneer axons in the optic stalk. RGC differentiation continues in centrifugal waves. Later born RGC axons fasciculate with the more mature axons. Growth cones at the end of the axons respond to guidance cues to adopt a centripetal direction, maintain nerve fiber layer restriction and to leave the optic cup. Although there is extensive information on OC-OS development, we still have important unanswered questions regarding its contribution to the exit of the RGC axons out of the eye. We are still to distinguish the morphogens of the OC-OS from the axon guidance molecules which are expressed in the same place at the same time. The early RGC transcription programs responsible for axon emergence and pathfinding are also unknown. This review summarizes the molecular mechanisms for early RGC axon guidance by contextualizing mouse knock-out studies on OC-OS development with the recent transcriptomic studies on developing RGCs in an attempt to contribute to the understanding of human optic nerve developmental anomalies. The published data summarized here suggests that the developing optic nerve head provides a physical channel (the closing optic fissure) as well as molecular guidance cues for the pioneer RGC axons to exit the eye

Morphologies of mouse retinal ganglion cells expressing transcription factors Brn3a, Brn3b, and Brn3c: Analysis of wild type and mutant cells using genetically-directed sparse labeling

AbstractThe mammalian retina contains more than 50 distinct neuronal types, which are broadly classified into several major classes: photoreceptor, bipolar, horizontal, amacrine, and ganglion cells. Although some of the developmental mechanisms involved in the differentiation of retinal ganglion cells (RGCs) are beginning to be understood, there is little information regarding the genetic and molecular determinants of the distinct morphologies of the 15–20 mammalian RGC cell types. Previous work has shown that the transcription factor Brn3b/Pou4f2 plays a major role in the development and survival of many RGCs. The roles of the closely related family members, Brn3a/Pou4f1 and Brn3c/Pou4f3 in RGC development are less clear. Using a genetically-directed method for sparse cell labeling and sparse conditional gene ablation in mice, we describe here the sets of RGC types in which each of the three Brn3/Pou4f transcription factors are expressed and the consequences of ablating these factors on the development of RGC morphologies

Postnatal developmental dynamics of cell type specification genes in Brn3a/Pou4f1 Retinal Ganglion Cells

Abstract Background About 20–30 distinct Retinal Ganglion Cell (RGC) types transmit visual information from the retina to the brain. The developmental mechanisms by which RGCs are specified are still largely unknown. Brn3a is a member of the Brn3/Pou4f transcription factor family, which contains key regulators of RGC postmitotic specification. In particular, Brn3a ablation results in the loss of RGCs with small, thick and dense dendritic arbors (‘midget-like’ RGCs), and morphological changes in other RGC subpopulations. To identify downstream molecular mechanisms underlying Brn3a effects on RGC numbers and morphology, our group recently performed a RNA deep sequencing screen for Brn3a transcriptional targets in mouse RGCs and identified 180 candidate transcripts. Methods We now focus on a subset of 28 candidate genes encoding potential cell type determinant proteins. We validate and further define their retinal expression profile at five postnatal developmental time points between birth and adult stage, using in situ hybridization (ISH), RT-PCR and fluorescent immunodetection (IIF). Results We find that a majority of candidate genes are enriched in the ganglion cell layer during early stages of postnatal development, but dynamically change their expression profile. We also document transcript-specific expression differences for two example candidates, using RT-PCR and ISH. Brn3a dependency could be confirmed by ISH and IIF only for a fraction of our candidates. Conclusions Amongst our candidate Brn3a target genes, a majority demonstrated ganglion cell layer specificity, however only around two thirds showed Brn3a dependency. Some were previously implicated in RGC type specification, while others have known physiological functions in RGCs. Only three genes were found to be consistently regulated by Brn3a throughout postnatal retina development – Mapk10, Tusc5 and Cdh4

Equipment for testing the indications accuracy of speedometers and altimeters existing on board aircraft and the tightness of the related pneumatic paths

The equipment is intended to testing the tightness of the catchment pneumatic system (Pitot tube), the transmission (pneumatic paths) and the total and static air pressures processing (aircrafttype instruments in order to establish the main flight parameters and checking the correctness of the operation of related aircraft instruments: the altimeter and the speedometer

Influence of Heat Treatment on the Corrosion Behavior of Electrodeposited CoCrFeMnNi High-Entropy Alloy Thin Films

In this paper, we investigate what effects heat treatment can have on potentiodynamically electrodeposited high-entropy thin film (HEA) CoCrFeMnNi alloys. We focused our study on the corrosion resistance in synthetic seawater, corroborated with the structure and microstructure of these thin films. Thin films of HEA alloys were deposited on a copper foil substrate, using an electrolyte based on the organic system dimethyl-sulfoxide (DMSO-(CH3)2SO)-acetonitrile (AN-CH3CN) (in a volume ratio of 4:1), which contains LiClO4 as electrolyte support and chloride salts of CoCl2, CrCl3 × 6H2O, FeCl2 × 4H2O, MnCl2 × 4H2O and NiCl2 × 6H2O. Using MatCalc PC software, based on the CALPHAD method, the structure and characteristics of the HEA system were investigated, and thermodynamic and kinetic criteria were calculated. The modeling process generated in the body-centered-cubic (BCC) or face-centered-cubic (FCC) structures a series of optimal compositions that are appropriate to be used in anticorrosive and tribological applications in a marine environment. Electrochemical measurements were carried out in an aerated artificial seawater solution at ambient temperature. In the experimental media, HEA thin films proved to have good corrosion resistance and were even better than the copper substrate. Corrosion resistance was improved after heat treatment, as shown by polarization and EIS tests. The structure and microstructure of HEA thin films before and after corrosion in artificial seawater were investigated by X-ray diffraction (XRD), scanning electron microscopy (SEM) and energy dispersive spectrometry (EDS). The XRD data showed no significant changes in the structure of HEA heat-treated thin films after the corrosion in saline media. The data obtained by polarization and ESI are supported by results from SEM-EDS. This complex study reveals that, for HEA thin films, heat treatment leads to an increase in corrosion resistance. So, this finding suggests that thermal annealing is an appropriate method for improving the corrosion performance of HEA thin films

Expression of Brn3a in the spinal cord at embryonic stages.

(A-D) Immunostaining of Brn3a (green) was performed on the transverse sections of the thoracic spinal cord from C57BL/6J mice at E12.5 (A), E14.5 (B), E16.5 (C), and E18.5 (D). The sections were counterstained by Hoechst 33342 (blue). Brn3a-void region in the dorsal horn is marked by square brackets. Spinal cords on the right side are shown. Arrows in D indicate Brn3a-positive neurons in the marginal region. Schematic diagrams of the spinal cord are shown in the insets. Red dotted squares indicate the area shown in the image. (E-J) Immunostaining of Brn3a (green), together with Brn3b (magenta), was performed on the transverse sections of the thoracic spinal cord from C57BL/6J mice at E12.5 (E-G) and E18.5 (H-J). Scale, 200 μm. (K) The number of Brn3a (red)- and Brn3b (blue)-positive cells in the hemi-spinal cord at E12.5 and E18.5 is shown. (L) The percentage of Brn3a- and Brn3b-double positive cells among Brn3a-positive cells at E12.5 and E18.5 is shown. Data are presented as the mean ± SEM.</p

Labeling of Brn3a-KO neurons by using Brn3a-cKOAP mice.

(A) pCAG-Cre together with pCAG-nlsEGFP were introduced into spinal dorsal horn neurons of Brn3a-cKOAP mice at E11.8 by in utero electroporation, and the spinal cord was dissected out at E18.5. AP assay was performed on the transverse section of the spinal cord to visualize the distribution of Brn3a-KO neurons. (B-D) AP signal (magenta; B, D) and nlsEGFP fluorescence (green; C, D) on the right spinal dorsal horn are shown. Arrows indicate double-positive neurons. Scale, 200 μm. (PDF)</p

Effect of Brn3a overexpression on the axonal extension of Brn3a-lineage neurons.

pCAG-LSL-EGFP with or without pCAG-Brn3a was introduced into the spinal dorsal horn neurons of Brn3aCre/+ mice, as shown in Fig 6. The number of EGFP-positive axons normalized by that of EGFP-positive cell bodies in each section was defined as the index. The index of control (n = 5) and Brn3a-overexpressing (n = 5) mice in the DF on the ipsilateral side (A) as well as in the VF and VLF on the ipsilateral (B) and contralateral (C) sides is shown. Significant differences were assessed using the Mann–Whitney U-test. (A) p = 0.6905. (B) p = 0.0079. (C) p = 0.0317. “ns” = non-significant. *p p < 0.01. Data are the mean ± SEM.</p

Axonal extension index of control and Brn3a-overexpressed neurons.

Axonal extension index of control and Brn3a-overexpressed neurons.</p